Interpretive Summary: Red imported fire ants, a mound-building ant species, are a major agricultural and urban pest throughout the southeastern United States. When building their nests, fire ants incorporate chemicals in the nest material. Knowledge of fire ant nest chemistry is important not only in understanding behavior, chemical communication and nest microclimate regulation, but also for the potential to exploit the ant chemical signals in fire ant management. A novel experimental technique was developed and a series of chemicals were identified in the nesting material. Some of these identified chemicals may be used as attractants and stabilizers in fire ant bait formulations.

Technical Abstract:
Red imported fire ants are believed to incorporate ant-derived chemicals in nesting material. However, only a few chemicals have been identified. One hurdle for such investigation is the interference in chemical analysis from soil-borne chemicals. Ants were found to be able to construct their nests using wetted silica gel as the only building material in the laboratory. This provided an opportunity to identify ant-derived chemicals in nest material without dealing with the interference from soil-borne chemicals. Ants were allowed to build their nest in a device using wetted silica gel as the only nesting material. A similar device with silica gel was used as a control in which no ants were released. Workers built nests by excavating silica gel and making honeycomb-like structures in the device. After building the nests, silica gel samples were collected, extracted with pentane, pyridine or water, and analyzed using gas chromatography - mass spectrometry (GC-MS). Since no food was provided during the experiment, any chemicals which existed in silica gel in the nesting device but not in the control were assumed to be from the ants. Ants seemed to prefer depositing their excreta on the edge of the nesting device, which were mixed with silica gel. Since excreta are believed to contribute to the chemical modification of ant nest soil in the field, excreta samples were also analyzed. All silica gel samples contained cuticular hydrocarbons. Venom alkaloids, phosphoric acid, glycerol, lactic acid, and malonic acid were identified in samples collected inside the nest. Uric acid, 2,6-dihydroxy-9H-purine, urea, glycerol, phosphoric acid, amino acids, and organic acids were found in fire ant excreta.